The main goal of the proposed research is to determine the structure of the principal neutralizing determinant (PND) of HIV-1 complexed with specific monoclonal antibodies. The HIV-1 PND maps to a disulfide- linked loop in the third variable domain, the V3 region of gp120. Antisera raised against this loop usually neutralize the virus in an isolate-restricted manner. However, recent results suggest it may be possible to achieve broader reactivity. The specific goals of this proposal are to determine the structures of the PND from different isolates. The antibodies to be studied are mouse 50.1 and 83.1 (MN specific), the more broadly neutralizing 59.1 and 58.2 (MN, IIIB, WMJ2, SF2) and human antibodies selected for binding to PND peptides and to designed cyclized peptides. The human antibodies will be selected for study from a comparison of their binding profiles with the antisera of infected patients to ensure biologically important PND conformations are being studied. It is then the intent to determine the conformation of the PND bound to each antibody and hence to understand how different antibodies recognize different isolates. The structure of the broadly neutralizing antibodies should provide invaluable insights into the broad recognition and neutralization of different HIV-l strains. A novel approach to crystallizing gpl20 is proposed by complexing it with the neutralizing Fabs. Valuable information will be obtained not only of the structure and function of gpl20 but also the structure of the PND in the intact antigen. The structural information obtained from all those studies can be used in the iterative design of immunogens constrained to adopt their biologically relevant conformations which could be of value either as vaccines or as drugs to inhibit important biological functions of the HIV- l virus, such as viral fusion. The PND structures can also be used to understand how single amino acid changes affect viral tropism and biological functions such as syncytia formation. The structures will be determined by x-ray crystallography using molecular replacement (MR) or multiple isomorphous replacement. A brief summary of the specific aims are: 1. Crystal Structures of Isolate-Specific Murine Anti-HIV-1 Fabs with Linear PND Peptides. 2. Crystal Structures of Broadly Neutralizing Murine Anti-HIV-1 Fabs with Linear PND Peptides. 3. Crystal Structures of Murine Fabs with Cyclic PND Peptides and V3- Loop Mimics. 4. Crystal Structures of Human Anti HIV-1 Fabs with Linear and Cyclic PND Peptides and V3-Loop Mimics. 5. Design of Cyclized Peptides and V3-Loop Mimics. 6. Crystallization of gp120-Fab Complexes.